Further examination of the restriction-modification system of Hemophilus strengthened our conclusion that type I restriction enzymes play a role in the cell in restricting unmodified phage DNA, but that they do not restrict transforming DNA. Some mutants defective in phage restriction-modification are deficient in producing successful recombinant molecules. The role of type II restriction enzymes in the cell is still unclear since they do not restrict unmodified DNA in vivo but can act on unmodified DNA in vitro. A type I restriction enzyme has been found in partially purified extracts of type f H. influenzae along with a type II enzyme that has a different specificity than endonucleases from other H, influenzae stereotypes. A very strong "non-specific" nuclease has also been found. In addition a nuclease specific for heterologous single-stranded DNA has been identified and a factor has been found that stimulates transformation with H. influenzae type f single-stranded DNA. We are also studying the property of some DNAs that affect their ability to transform as single-stranded DNA. Studies also continue on the ability of some methylases to methylate their own DNA. Lastly we continue to investigate the instability of the genetic determinants of restriction and modification in H. influenzae type d. BIBLIOGRAPHIC REFERENCES: Roszczyk, Emilia and Goodgal, Sol H. Methylase activities from Haemophilus influenzae that protect H.parainfluenzae transforming DNA from inactivation by H.influenzae endo R. Journal of Bacteriology 123:287-293, 1975. Gromokova, Rosa and Goodgal, Sol H. The biological properties of a Haemophilus influenzae restriction enzyme Hind I. Submitted to the Journal of Bacteriology, 1975.